The present invention relates to novel cyclic peptides and salts thereof which have a VLA-4 adhesion molecule inhibition action useful as a therapeutic agent for inflammatory diseases, for example allergic inflammatory diseases such as bronchial asthma, atopic dermatitis and allergic rhinitis, hepatitis, nephritis, chronic rheumatoid arthritis, autoimmune disease, rejection reactions following organ transplantation, type I diabetes, Crohn""s disease, or postoperative restenosisis prevention and arteriosclerosis, and to medicinal uses thereof.
VLA-4 (very late antigen-4) is an adhesion molecule which is expressed on white blood cells such as lymphocytes, monocytes, basophils and eosinophils, and it belongs to the VLA family. VLA-4 has a heterodimer structure comprising an xcex1 chain and xcex2 chain, and is also referred to as xcex14xcex21, or CD49d/CD29. It plays a part in cell-cell and cell-extracellular matrix interactions, and is involved in the infiltration of white blood cells into sites of inflammation. VCAM-1 (vascular cell adhesion molecule-1) which is present on vascular endothelial cells and fibronectin of the extracellular matrix is known as an adhesion molecules which interact with VLA-4.
The binding site for VLA-4 on fibronectin is a fibronectin fragment referred to as CS-1 and, within this fragment, the smallest unit of amino acids necessary for binding is reported to be the three amino acid residues leucine-aspartic acid-valine. Furthermore, chain form or cyclic peptide VLA-4 adhesion inhibiting compounds based on the three amino acid residues leucine-aspartic acid-valine have been reported (WO95/15973).
On the other hand, with regard to VCAM-1, which is the other adhesion molecule which interacts with VLA-4, it is known that its manifestation mainly on vascular endothelial cells is increased by stimulation by cytokines such as IL-1, TNF-xcex1 and IL-4, and that it interacts with the VLA-4 present on cells such as lymphocytes, NK cells, monocytes andfeosinophils. VLA-4 and VCAM-1 play a part in the process:of white blood cell infiltration into inflammation sites from blood vessels and, from this point of view, the interaction between VLA-4 and VCAM-1 in the inflammatory response is extremely important.
Of the adhesion molecules, VCAM-1 belongs to an immunoglobulin super family, and 7-Ig-like-domain VCAM-1 and 6-Ig-like-domain VCAM-1 are known. Binding sites for VLA-4 on VCAM-1 are present at domain 1 and domain 4 and it has been made clear from the results of VCAM-1 mutation that, in particular within these domains, glutamine-isoleucine-aspartic acid-serine-proline, which is the amino acid sequence on the CD loop, is important in the binding with VLA-4 (J. Cell Bio., 125, 1395 (1994), J. Cell Bio., 124, 601 (1994), J. Cell Bio., 125, 215 (1994) and J. Cell. Science, 107, 2127 (1994)). Furthermore, taking glutamine-isoleucine-aspartic acid-serine-proline as the basic peptide, J. H. Wang et al have reported the cyclic peptide Cys* Gln Ile Asp Ser Pro Cys* SEQ ID NO:58 (where Cys*Cys* indicates a disulphide bond) which has a VLA-4 adhesion inhibiting activity (Proc. Natl. Acad. Sci. USA, 92, 5714 (1995)). Again, inhibitors which were designed making reference to the anti-VLAxcex14 antibody CDR3 amino acid sequence and the VCAM-1 EF loop amino acid sequence.
It has been made clear from animal models, specifically contact hypersensitivity and delayed hypersensitivity models (mouse and rat), experimental autoimmune encephalomyelitis models (mouse and rat), nephrotic nephritis models (rat), passive cutaneous anaphylaxis models (guinea pig), immune complex-induced lung injury models (rat), spontaneous colitis models (monkey) and asthma models (sheep), where anti-VLA-4 antibodies have been used, that VLA-4 plays an important role in the inflammatory response.
Conventional drugs which are used against inflammatory diseases such as allergic inflammation and chronic rheumatoid arthritis possess the action of inhibiting the action of chemical transmitters, or the action of suppressing the production of chemical transmitters or the action of inhibiting the production of active oxygen, etc, but since they do not have, as their main action, the action of suppressing the process of white blood cell infiltration to the inflammation site, they cannot suppress the progress of inflammation. As stated earlier, the adhesion molecules VLA-4 and VCAM-1 contribute mainly to the process of white blood cell infiltration to the inflammation site and so novel cyclic peptides which have the activity of inhibiting the adhesion of VLA-4 and VCAM-1 may be expected to suppress white blood cell infiltration and may be considered to have considerable potential as therapeutic agents effective against the aforesaid diseases. Hitherto, there have been reported inhibiting compounds synthesized with reference to CS-1, which is the binding site for VLA-4 on the fibronectin side, and the anti-VLAxcex14 antibody epitope, but except for Wang et al, there have been no reports of novel peptide compounds created with reference to the binding site on the VCAM-1 side, and to the medicinal applications thereof.
The objective of the present invention lies in creating compounds with reference to the binding site on the VCAM-1 side, and offering novel peptides.
As a result of painstaking investigations with reference to the binding site on the VCAM-1 side, the present inventors discovered that, by creating novel cyclic peptides with a marked VCA-4 adhesion inhibition activity, these cyclic peptides can be used as therapeutic agents for, amongst inflammatory diseases, allergic inflammatory diseases and hepatitis, and the a present invention has been perfected based on this discovery.
Specifically, the present invention, has the following constitution.
It relates to cyclic peptides represented by formula (I) 
[where, in formula (I),
A and F may be the same or different, and are both selected from the group comprising; L- or D-form Cys, homo-Cys, Pen and Mpr, or both from the group comprising Asp, Glu, Aad, Dpr, Dab and Orn,
B is selected from the group comprising L- or D-form Ala, Ala(t-Bu), Val, Leu, Ile, aIle, Abu, Nle, Nva, Tle, Cha, Chg, Phe, Phg, Trp, Ala(3-Bzt), Ala(1-Naph), Ala(2-Naph), Ala(2-Pyr), Ala(2-Qui), His, Thi, Ala(4-Thz), 2-Abz, Pro, homo-Pro and Tic,
C is selected from the group comprising L-form Asp analogues, Glu analogues, Aad analogues, Asn analogues, Gln analogues, Ser, Ser(OMe), homo-Ser, Dpr, Dab, Orn, Met, Met(O), Met(O2), aIle, Nle, Nva, Chg, Phg, Tyr and Tle and, when C is selected from L-form Asp analogues, Glu analogues and Aad analogues, then it is of formula (II) 
n=1 Asp n=2 Glu n=3 Aad
(where R3 may be hydrogen or an alkyl group and, when R3 is an alkyl group, it comprises a C1 to C6 straight chain or branched alkyl group) and,
when C is selected from.Asn analogues and Gln analogues, then it is of formula (III) 
n=1 Asn n=2 Gln
(where R4 and R5 may be the same or different, and may respectively be hydrogen or an alkyl group and, in the case where R4 or R5 is an alkyl group, it comprises a C1 to C6 straight chain or branched alkyl group),
D is selected from the group comprising L- or D-form Tyr, Ser, homo-Ser, Leu, Ile, aIle, Nle, Nva, Chg, Cha, Val, Ala(t-Bu), Abu, Tle, Ala, Phg, homo-Phe, Phe, Ala(2-Naph), Ala(2-Pyr), Ala(3-Bzt), Ala(1-Naph), Ala(2-Qui), Thi, Ala(4-Thz), 2-Abz, Trp and His, E is selected from the group comprising L- or D-form Leu, Ile, aIle, Nle, Nva, Chg, Cha, Tle, Phg, homo-Phe, Ala(2-Naph), Ala(2-Pyr), Ala(3-Bzt), Ala(1-Naph), Ala(2-Qui), Thi, Ala(4-Thz), 2-Abz and His, G represents a, disulphide bond or amide bond,
R1 may be hydrogen or an acyl group, and in the case where R1 is an acyl group, then it is. of formula (IV) 
(where R6 comprises a C1 to C6 straight chain or branched alkyl group, a C4 to C8 cycloalkyl group, a C5 to C11 alkylcycloalkyl group, a C6 to C14 aryl group or a C7 to C11 aralkyl group), and
R2 may be hydrogen or an alkyl group and, in the case where R2 is an alkyl group, it is a C1 to C6 straight chain or branched alkyl group],
or the salts thereof, and to medicinal applications thereof.